The present invention relates to novel compounds which have hemoregulatory activities and can be used to stimulate hematopoiesis and for the treatment of viral, fungal and bacterial infectious diseases.

Claim:

What is claimed is:

1. A method of treating viral, fungal or bacterial infections which comprises administering to an animal in need thereof, an effective amount of a compound of Formula (I):##STR3##

R.sup.2 is hydrogen, C.sub.1-4 alkylC(O)R.sub.4, C.sub.1-4 alkyl or R.sup.2 is benzyl which is optionally substituted by one or two C.sub.1-4 alkyl, C.sub.1-4 alkoxy, F, Cl, I, Br, OH, or N(R.sub.3).sub.2 ;

R.sub.3 is independently hydrogen, C.sub.1-4 alkyl, or benzyl;

R.sub.4 is independently OR.sub.3, N(R.sub.3).sub.2 or SR.sub.3 ; and

k, is an integer from 0 to 4;

m is 2;

n is independently 2 or 3;

y is zero or one;

or a pharmaceutically acceptable salt thereof.

2. A method of treatment according to claim 1 wherein the compound is selected from N,N'-bis(picolinoyl)piperazine, or N,N'-bis(pyroglutamoyl)piperazine.

3. A method of treating sepsis which comprises administering to an animal in need thereof, an effective amount of a compound of Formula (I): ##STR4##

R.sup.2 is hydrogen, C.sub.1-4 alkylC(O)R.sub.4, C.sub.1-4 alkyl or R.sup.2 is benzyl which is optionally substituted by one or two C.sub.1-4 alkyl, C.sub.1-4 alkoxy, F, Cl, I, Br, OH, or N(R.sub.3).sub.2 ;

R.sub.3 is independently hydrogen, C.sub.1-4 alkyl, or benzyl,

R.sub.4 is independently OR.sub.3, N(R.sub.3).sub.2 or SR.sub.3, and

k is an integer from 0 to 4;

m is 2;

n is independently 2 or 3;

y is zero or one;

or a pharmaceutically acceptable salt thereof.

Description:

FIELD OF THE INVENTION

The present invention relates to novel compounds which have hemoregulatory activities and can be used to stimulate hematopoiesis and for the treatment of viral, fungal and bacterial infectious diseases.

BACKGROUND OF THE INVENTION

The hematopoietic system is a life-long cell renewal process whereby a defined stem cell population gives rise to a larger population of mature, differentiated blood cells (Dexter T M. Stem cells in normal growth and disease. Br Med J 1987;195:1192-1194) of at least nine different cell lineages (erythrocytes, platelets, eosinophils, basophils, neutrophils, monocytes/macrophages, osteoclasts, and lymphocytes) (Metcalf D. The Molecular Control of Blood Cells. 1988; Harvard University Press,Cambridge, Mass.). Stem cells are also ultimately responsible for regenerating bone marrow following treatment with cytotoxic agents or following bone marrow transplantation.

The major dose-limiting toxicities of most standard anti-neoplastic drugs are related to bone marrow suppression, which if severe and prolonged, can give rise to life-threatening infectious and hemorrhagic complications. Myelosuppression ispredictable and has been reported to be dose-limiting in greater than 50% of single-agent Phase I trials cytotoxic compounds (Merrouche Y, Catimel G, Clavel M. Hematopoietic growth factors and chemoprotectants; should we move toward a two-step processfor phase I clinical trials in oncology? Ann Oncol 1993; 4:471-474). The risk of infection is directly related to the degree of myelosuppression as measured by the severity and duration of neutropenia (Brody G P, Buckley M, Sathe Y S, Freireich E J.Quantitative relationship between circulating leukocytes and infections with acute leukemia. Ann In Med 1965; 64:328-334).

The control of hematopoiesis involves the interplay of a variety of cytokines and growth factors during various stages of the hematopoietic cascade, including early pluripotent stem cells and mature circulating effector cells. These regulatorymolecules include granulocyte colony stimulating factor (G-CSF), granulocyte-macrophage stimulating factor (GM-CSF), macrophage-colony stimulating factor (M-CSF), and a variety of interleukins which have overlapping, additive and synergistic actionswhich play major roles in host defence. Mechanistically, this is accomplished by enhancing the production of granulocytes and macrophages, as well as by the activation of effector cell functions (Moore M A S. Hemopoietic growth factor interactions: invitro and in vivo preclinical evaluation. Cancer Surveys 1990; 9:7-80). These coordinated activities support optimal host defences which are necessary for fighting bacterial, viral and fungal infections.

Strategies to prevent or reduce the severity of neutropenia and myelotoxicity include the use of hematopoietic growth factors and/or other hematopoietic cytokines. Such treatments are becoming common practice, in that they offer the potential ofincreased doses of cytotoxic agents that may improve the therapeutic efficacy of antineoplastic agents, and reduce the morbidity associated with their use (Steward W P. Granulocyte and granulocyte-macrophage colony stimulating factors, Lancet 1993;342:153-157). Clinical studies have demonstrated the G-, GM- and/or M-CSF may reduce the duration of neutropenia, accelerate myeloid recovery, and reduce neutropenia-associated infections and other infectious complications in patients with malignancieswho are receiving cytotoxic chemotherapy or in high infectious-risk patients following bone marrow transplantation (Steward W P. Granulocyte and granulocyte-macrophage colony stimulating factors, Lancet 1993; 342:153-157 and Munn D H, Cheung N K V.Preclinical and clinical studies of macrophage colong-stimulating factor. Semin Oncol 1992; 19:395-407).

Synthetic peptides have been reported to induce the synthesis and release of haematoporetic mediators, including m-CSF from bone marrow stromal elements see U.S. patent application Ser. No. 08/001,905.

We have now found certain novel non-peptide compounds which have a stimulative effect on myelopoietic cells. They are useful in stimulating myelopoiesis in patients suffering from reduced myelopoietic activity, including bone marrow damage,agranulocytosis and aplastic anemia including patients having depressed bone maprow function due to immunosuppressive treatment to suppress tissue reactions i.e. in bone marrow transplant surgery. They may also be used to promote more rapid regenerationof bone marrow after cytostatic chemotherapy and radiation therapy for neoplastic an d viral diseases. They may be of particular value where patients have serious infections due to a lack of immune response following b one marrow failure . They areuseful in the treatment and pievention of viral, fungal and bacterial disease.

SUMMARY OF THE INVENTION

This invention comprises compounds, hereinafter represented as Formula (I), which have hemoregulatory activities and can be used to stimulate hematopoiesis and in the prevention and treatment of bacterial, viral and fungal diseases.

These compounds are useful in the restoration of leukocytes in patients with lowered cell counts resulting from a variety of clinical situations, such as surgical induced myclosuppression, AIDS, ARDS, congenital myelodysplacis, bone marrow andorgan transplants; in the protection of patients with leukopenia from infection; in the treatment of severely burned patients and in the amelioration of the myelosuppression observed with some cell-cycle specific antiviral agents and in the treatment ofinfections in patients who have had bone marrow trasplants, especially those with graft versus host disease, in the treatment of tuberculosis and in the treatment of fevers of unknown origin in humans and animals. The compounds are also useful in thetreatment and prevention of viral, fungal and bacterial infectious diseases, particularly Candida and Herpes in both immunosuppressed and "normal" subjects. They are useful in the treatment of sepsis caused by gram negative and gram positive organisms.

These compounds may also be used in combination with the myelosuppresive agents of co-pending U.S. application Ser. No. 07/99,465 and U.S. Pat. No. 4,499,081, incorporated by reference herein, to provide alternating peaks of high and lowactivity in bone marrow cells, thus augmenting the natural circadian rhythm of hematopoiesis. In this way, cytostatic therapy can be given at periods of low bone marrow activity, thus reducing the risk of bone marrow damage, while regeneration will bepromoted by the succeeding peak of activity. This invention is also a pharmaceutical composition, which comprises a compound of Formula (I) and a pharmaceutically acceptable carrier.

This invention further constitutes a method for stimulating the myelopoietic system of an animal, including humans, which comprises administering to an animal in need thereof, an effective amount of a compound of Formula (I).

This invention also constitutes a method for preventing and treating viral, fungal and bacterial infections in immunosuppressed and normal animals, including humans, which comprises administering to an animal in need thereof, an effective amountof a compound of Formula (I).

DETAILED DESCRIPTION OF THE INVENTION

The compounds of the invention are represented by structural Formula I ##STR1##

Z is independently a 4-10 membered mono- or bicyclic heterocyclic ring system containing up to four heteroatoms N, O, S in the ring in which at least one heteroatom is N, and wherein the ring is substituted or unsubstituted by one or twoC.sub.1-4 alkyl, F, Cl, Br, I, C.sub.1-4 alkoxy, (CH.sub.2).sub.m R.sub.4, oxo, oxime, O--C.sub.1-4 alkyloxime, hydroxy, N(R.sub.3).sub.2, acylamino or aminoacyl groups, 8, 9, 10 membered monocyclic ring systems being excluded;

R.sup.1 and R.sup.2 are independently hydrogen, C.sub.1-4 alkylC(O)R.sub.4, C.sub.1-4 alkyl or R.sub.1 and R.sub.2 are benzyl which is optionally substituted by one or two C.sub.1-4 alkyl, C.sub.1-4 alkoxy, F, Cl, I, Br, OH, or N(R.sub.3).sub.2 ;

R.sub.3 is independently hydrogen, C.sub.1-4 alkyl, or benzyl;

R.sub.4 is independently OR.sub.3, N(R.sub.3).sub.2 or SR.sub.3 ; and

k is an integer from 0 to 4;

m is independently an integer from 1 to 3;

n is 1 or 2;

y is zero or one;

or a pharmaceutically acceptable salt thereof.

C.sub.1-4 alkyl groups may be straight or branched.

The compounds of the present inventiion may contain one or more asymmetric carbon atoms and may exist in racemic and optically active form. All these compounds and diastereomers are contemplated to be within the scope of the present invention.

Compounds of Formula (I) wherein A.sup.1, A.sup.2 and m are defined as in Formula (I) are prepared by methods analogous to those described in Scheme 3. Appropriate diamines (such as 1 in Scheme 3) are bis-acylated with a suitable heterocyclicacid (such as 2 in Scheme 3) using an activating agent (such as BOP reagent) with a suitable base (such as iPr.sub.2 NEt) in a polar aprotic solvent (such as DMF) to give final product. ##STR2##

In order to use a compound of the Formula (I) or a pharmaceutically acceptable salt thereof for the treatment of humans and other mammals it is normally formulated in accordance with standard pharmaceutical practice as a pharmaceuticalcomposition.

According to a still further feature of the present invention there are provided pharmaceutical compositions comprising as active ingredient one or more compounds of Formula (I) as herein before defined or physiologically compatible saltsthereof, in association with a pharmaceutical carrier or excipient. The compositions according to the invention may be presented for example, in a form suitable for oral, nasal, parenteral or rectal administration.

As used herein, the term "pharmaceutical" includes veterinary applications of the invention. These compounds may be encapsulated, tableted or prepared in an emulsion or syrup for oral administration. Pharmaceutically acceptable solid or liquidcarriers may be added to enhance or stabilize the composition, or to facilitate preparation of the composition. Liquid carriers include syrup, peanut oil, olive oil, glycerin, saline and water. Solid carriers include starch, lactose, calcium sulfatedihydrate, terra alba, magnesium stearate or stearic acid, talc, pectin, acacia, agar or gelatin. The carrier may also include a sustained release material such a glyceryl monostearate or glyceryl distearate, alone or with a wax. The amount of solidcarrier varies but, preferably will be between about 20 mg to about 1 g per dosage unit. The pharmaceutical preparations are made following the conventional techniques of pharmacy involving milling, mixing, granulating, and compressing, when necessary,for tablet forms; or milling, mixing and filling for hard gelatin capsule forms. Capsules containing one or several active ingredients may be produced, for example, by mixing the active ingredients with inert carriers, such as lactose or sorbitol, andfilling the mixture into gelatin capsules. When a liquid carrier is used, the preparation will be in the form of a syrup, elixir, emulsion or an aqueous or non-aqueous suspension. Such a liquid formulation may be administered directly p.o. or filledinto a soft gelatin capsule. Organ specific carrier systems may also be used.

Alternately pharmaceutical compositions of the compounds of this invention, or derivatives thereof, may be formulated as solutions of lyophilized powders for parenteral administration. Powders may be reconstituted by addition of a suitablediluent or other pharmaceutically acceptable carrier prior to use. The liquid formulation is generally a buffered, isotonic, aqueous solution. Examples of suitable diluents are normal isotonic saline solution, standard 5% dextrose in water or bufferedsodium or ammonium acetate solution. Such formulation is especially suitable for parenteral administration, but may also be used for oral administration and contained in a metered dose inhaler or nebulizer for insufflation. It may be desirable to addexcipients such as polyvinylpyrrolidone, gelatin, hydroxy cellulose, acacia, polyethylene glycol, mannitol, sodium chloride or sodium citrate.

For rectal administration, a pulverized powder of the compounds of this invention may be combined with excipients such as cocoa butter, glycerin, gelatin or polyethylene glycols and molded into a suppository. The pulverized powders may also becompounded with an oily preparation, gel, cream or emulsion, buffered or unbuffered, and administered through a transdermal patch.

Nasal sprays may be formulated similarly in aqueous solution and packed into spray containers either with an aerosol propellant or provided with means for manual compression.

Dosage units containing the compounds of this invention preferably contain 0.05-50 mg, for example 0.05-5 mg of the compound of formula (V) or salt thereof.

According to a still further feature of the present invention there is provided a method of stimulation of myelopoiesis which comprises administering an effective amount of a pharmaceutical composition as hereinbefore defined to a subject.

No unacceptable toxicological effects are expected when compounds of the invention are administered in accordance with the present invention.

The biological activity of the compounds of Formula (I) are demonstrated by the following tests.

The superoxide dismutase-inhibitable (SOD) superoxide released by effector cells in response to a in vitro stimulation by phorbol myristate acetate (PMA) (100-200 nM) or pre-opsonized (autologous sera) live C. albicans (E:T=1:10) are quantitatedin a microtiter ferricytochrome c reduction assay. The assay is performed in the presence of 1% gelatin/HBSS and 80 uM ferricytochrome c in a total volume of 200 uL/well. The nmoles of cytochrome c reduced/well is calculated from spectrophotometricreadings (550 nm) taken following a 1 hour incubation at 37.degree. C. (5% CO.sub.2). The amount of SOD-inhibitable cytochrome c reduced is determined by the inclusion of wells containing SOD (200 U/well). Baseline superoxide release is determined inthe absence of stimuli. Experimental data are expressed as a percentage of the control group.

The following examples are illustrative and are not limiting of the compounds of this invention.

EXAMPLE 1

N,N'-Bis(picolinoyl)-1,4-diazacycloheptane

To a solution of 1,4-diazacycloheptane (0.31 g, 3.08 mmol) in pyridine (30 mL) was added picolinic acid (0.74 g, 6.01 mmol) and EDC (1.16 g, 6.05 mmol). After 18 h at room temperature, the bulk of the pyridine was removed in vacuo. The residuewas dissolved in water (50 mL) and extracted with EtOAc (3.times.50 mL). The combined organic extracts were washed with water (50 mL) and dried over Na.sub.2 SO.sub.4. Removal of solvent gave 0.14 g of a red syrup. Purification by flash chromatography(20 % MeOH/EtOAc, silica gel) afforded 0.09 g (10%) of the title compound. MS (ES+) m/z 311.2 [M+H].sup.+.

Step 2 Add sufficient water portion-wise to the blend from Step 1 with careful mixing after each addition. Such additions of water and mixing until the mass is of a consistency to permit its converion to wet granules.

Step 3 The wet mass is converted to granules by passing it through an oscillating granulator using a No. 8 mesh (2.38 mm) screen.

Step 4 The wet granules are then dried in an oven at 140.degree. F. (60.degree. C.) until dry.

Step 5 The dry granules are lubricated with ingredient No. 5.

Step 6 The lubricated granules are compressed on a suitable tablet press.

Parenteral Formulation

A pharmaceutical composition for parenteral administration is prepared by dissolving an appropriate amount of a compound of Formula I in polyethylene glycol with heating. This solution is then diluted with water for injections Ph Eur. (to 100ml). The solution is then sterilized by filtration through a 0.22 micron membrane filter and sealed in sterile containers.